The mechanisms that control the cell cycle are remarkably conserved among all eukaryotes. Both mammals and budding yeast commit to another round of cell division during G1. Oncogenic processes exert their greatest impact by interfering with regulators of G1 progression. We propose to investigate the molecular mechanisms which control G1 progression in budding yeast. Our experiments are designed to determine how commitment to the budding yeast cell cycle is regulated in response to nutritional shifts, cell growth and adversities, such as DNA damage. In yeast, as in all higher eukaryotes, cell cycle transitions are governed by cyclin-dependent kinases (Cdks). Nine cyclins have been identified that bind and activate the Cdk of budding yeast (Cdc28). Three cyclins Cln1, Cln2 and Cln3 play critical roles in modulating the G1 to S transition. Our focus is upon determining what controls the expression of these three cyclins during G1. ECB elements activate transcription of CLN3 and other key cell cycle regulators at the M/G1 boundary. Two other promoter elements have been identified that are activated during G1 by Swi4/Swi6 and/or Mbpl/Swi6. These complexes activate transcription of many genes including CLN1 and CLN2, which are rate limiting for the transition to S phase. Our goal is to understand how internal and external signals modulate the activity of these transcription complexes and control the transition to S phase. We have identified one activator and two repressors that influence ECB activity. We will use genetic and biochemical strategies to determine how cell cycle regulation of transcription is conferred by these elements. We will also investigate the regulatory role of Swi6 in activating transcription in G1 and inhibiting transcription in response to DNA damage. We have identified sites at which Rad53 kinase phosphorylates Swi6 in response to DNA damage. We will determine the significance of these phosphorylations to the G1 DNA damage checkpoint. We will also determine whether the consensus site for Rad53 phosphorylation that we draw from our studies enables us to predict the sites at which Rad53 modifies other known targets.